Previous studies have demonstrated that exogenous insulin, injected into central cerebrospinal fluid cavities of dogs, stimulates the release of endogenous insulin from the pancreas. To determine whether this response was elicited by (1) insulin per se, (2) an effect of insulin on glucose transport, or (3) glucopenia in the cerebrospinal fluid, we measured plasma insulin, glucose, and free fatty acids during the infusion of insulin, glucose, and 2-deoxy-glucose (2DG), individually or in combination, into the third cerebral ventricle of conscious dogs. As expected, the third ventricular infusion of insulin alone elicited a small, but significant, rise of plasma insulin. Surprisingly, infusion of insulin with glucose produced a smaller increase of plasma insulin (P less than 0.05) and the infusion of insulin with 2DG produced a much larger increase of plasma insulin (P less than 0.05) than did the third ventricular infusion of insulin alone. The third ventricular infusion of either glucose alone or 2DG alone had no effect on the plasma levels of insulin. These data suggest that administration of insulin into the cerebral ventricles stimulates pancreatic insulin secretion but not by accelerating the transport of glucose into a chemosensitive area of the brain.